The long-range objective of the proposed research is an understanding of the molecular basis of the pharmacological effects of narcotic analgesics, including analgesia and the development of tolerance and physical dependence. The recent discovery of stereospecific opiate receptors and endogenous opioid peptides (endorphins) in animal and human brain have opened research approaches that should aid in the attainment of this goal. Our research will be concerned with the further characterization of the opiate receptors and the elucidation of the functions of the endorphins. The interaction between the receptors and opiate agonists, antagonists and endorphins will be studied with special emphasis on the question of the existence of multiple types of receptors and on the influence of environmental and physiological factors on receptor number and properties. Physical chemical approaches will probe the conformation of opiate receptors and the surrounding membrane following opioid ligand binding and the mechanism of the sodium effect. Efforts to concentrate membrane-bound receptors and to solubilize and purify opiate receptors will be continued with special emphasis on the design of useful specific photoaffinity labeling agents and effective affinity chromatographic procedures. Assays for total endorphins and sensitive radioimmunoassays for individual opioid peptides in tissue extracts and fluids will be developed, as will methods for peptide separation and purification. These methods will be used to study endorphin levels and turnover in a variety of environmental, physiological and pathological conditions. It is hoped that such studies will aid in the understanding of the physiological function of endorphins. Organotypic tissue cultures of mouse fetal spinal cord with attached dorsal root ganglia will be used for studies of opiate receptors and endorphins in a system that permits manipulation of environmental factors as well as correlation of biochemical and electrophysiological changes.